Susceptibility of Pseudomonas aeruginosa to Carbenicillin

Ninety clinical isolates of Pseudomonas aeruginosa were examined for susceptibility to carbenicillin by the broth dilution and disc diffusion methods. Inhibition zone diameters varied at given minimal inhibitory concentration levels of the antibiotic. Nevertheless, the results obtained allowed the proposal of the following tentative criteria for the interpretation of inhibition zones. Pseudomonadaceae yielding zones of inhibition measuring at least 10 and 16 mm in diameter around 25-and 100-mug discs, respectively, are sensitive to this antibiotic when examined by the standardized Bauer-Kirby method of disc susceptibility testing. Isolates characterized by zones of less than 100 mm in diameter around 25-mug discs should be tested with 100-mug discs before they are reported as sensitive or resistant to carbenicillin.     

A new screening method for investigating microbial interactions

Interactions among Candida albicans, Staphylococcus aureus and Escherichia coli were investigated using a screening system in which test micro-organisms were incorporated in agar discs and effector micro-organisms in fluid growth media. Total as well as partial inhibition of test micro-organisms was observed in agar discs when these were incubated in broths containing effector micro-organisms. The ratio of numbers of test to effector micro-organisms was found to be of importance in the inhibition effect. The technique was found to be cheap, simple and versatile.     

Development of a method for the quantitative assessment of microorganism sensitivity to antibiotics using discs. The study of different nutrient media for determining microorganism sensitivity to antibiotics

Five nutrient media used for determination of microbial sensitivity to antibiotics, i.e. beaf-peptone agar, Hottinger pancreatic beaf infusion agar, sprat hydrolysate nutrient agar of the Dagestan Research Institute of Nutrient Media, Muller-Chintone agar from Bulgaria and "Oxoid" agar for determination of microbial sensitivity were studied comparatively. The media were compared with respect to the growth density with the use of different test-cultures and the clearance of the inhibition growth zones around the discs containing different antibiotics. The best results were obtained with the use of sprat hydrolysate nutrient agar. Further studies on the medium standardization are necessary.     

Properties of lipid-apolipoprotein association products. Complexes of human apo AI and binary phospholipid mixtures

The products resulting from the association of human apo A-I with binary phospholipid mixtures of dimyristoyl phosphatidylcholine (DMPC) and either dipalmitoyl (DPPC) or distearoyl (DSPC) phosphatidylcholine have been isolated and characterized. Effective lipid . protein complex formation was found to occur at the onset temperature for melting of the gel state, and equal incorporation of both lipid components of the binary mixture was observed. Two sizes of products were obtained, one containing 2 A-I molecules per complex and the other containing 3; the proportions of these two products depended upon the initial phospholipid/protein ratio employed. these two product species were found to be resolvable by density gradient centrifugation as well as gel filtration, reflecting substantial differences in composition as well as size. The ratio of DMPC to DPPC or DSPC was the same in the isolated complexes as in the initial mixture, suggesting that th protein does not associate preferentially with the fluid phase lipid, but with lipid domains in which the components are randomly distributed. Electron microscopy of recombinant particles containing a 2:1 ratio (w/w) of DSPC to DMPC revealed stacks of discs whose thickness was proportionately greater than for discs containing DMPC alone. Also of significance was the finding that recombinant discs containing 3 A-I molecules possessed a diameter approximately 1.5 times larger than recombinant discs containing 2 A-I molecules. These data are consistent with the model for the recombinant particles described by Tall et al. (Tall, A.R., Small, D.M., Deckelbau, R.J., and Shipley, G.G. (1977) J. Biol. Chem. 252; 4701-4711), in which the phospholipid is found as a circular bilayer, the thickness of which is dependent upon the length of the acyl chain, and around which the protein is distributed as an annulus.     

Ethylene-controlled Induction of Phenylalanine Ammonia-lyase in Citrus Fruit Peel

l-Phenylalanine ammonia-lyase (PAL) activity is low in the external layers (flavedo) of intact mature grapefruit peel. Flavedo discs evince upon incubation increasing PAL activity and ethylene production. Light has no effect in enhancing PAL activity in discs. Exogenous ethylene stimulates PAL activity in the flavedo of intact mature grapefruits (half maximum stimulation at 15 ppm); such activity rapidly decreases when fruit is removed from the ethylene containing atmosphere. Carbon dioxide inhibits both ethylene production and PAL activity of discs; exogenous ethylene only partly relieves PAL inhibition. Cycloheximide inhibits both PAL activity and ethylene production by flavedo discs. The same concentration of cycloheximide also inhibits PAL activity of discs in the presence of exogenous ethylene. Protein synthesis seems therefore to be needed at both levels of ethylene evolution and enhancement of PAL activity.     

Interpretation of Diffusion Susceptibility Data Obtained with 50-μg Carbenicillin Discs Against Gram-Negative Organisms

A total of 284 clinical isolates of various species of Enterobacteriaceae, Pseudomonas aeruginosa, P. maltophilia, and Acinetobacter anitratum were tested for susceptibility to carbenicillin by the standardized Bauer-Kirby disc diffusion technique and a microtiter broth dilution method. The data obtained led to the following proposed criteria for the interpretation of the results of disc susceptibility tests. Enterobacteriaceae that yield zones of inhibition equal to or greater than 20 mm in diameter around 50-mug discs of carbenicillin are designated as sensitive to the drug; isolates that yield zones measuring from 18 to 19 mm in diameter are reported as of equivocal (intermediate) susceptibility to the drug, whereas those enterobacterial isolates that are characterized by zones of inhibition of 17 mm or less in diameter are interpreted as resistant to carbenicillin. Isolates of P. aeruginosa, P. maltophilia, and A. anitratum yielding zones of 14 mm or more in diameter around 50-mug discs of carbenicillin are reported as sensitive, whereas those isolates that are characterized by zones of 13 mm or less in diameter are reported as resistant to this drug.     

双平板对照筛选跟踪分离细菌外排泵抑制剂

建立了细菌外排泵抑制剂的筛选与活性跟踪方法.准备2个平板,一个为普通营养琼脂平板,另一个为舍小蘖碱的普通营养琼脂平板,通过比较两个平板含药纸片周围抑菌圈的直径大小判断筛选结果,方法可靠稳定.筛选发现某霉菌提取物对细菌外排泵有抑制活性,经活性跟踪分离,得到单体化合物,经NMR鉴定为4′,5,7-三羟基异黄酮.方法简便易行,成本低,适宜于对大批样本进行快速筛选并在分离时进行活性跟踪.     

Effect of Antibiotics on Staphylococcal Hemolysin Production

Thirty-seven per cent of 126 strains of Staphylococcus aureus, when tested on sheep blood-agar with sensitivity discs containing cephalothin, carbenicillin, oxacillin, penicillin, and cycloserine, produced rings of beta-hemolysis surrounding zones of inhibition of bacterial growth. Each strain capable of producing a ring of beta-hemolysis did so with at least two of the mentioned antibiotics. None of the other 15 antibiotics tested was associated with a ring of beta-hemolysis surrounding any of the zones of inhibition. It appears, therefore, that the beta-hemolysis observed was produced by certain strains of S. aureus only in association with certain antibiotics.     

Metal resistance among aerobic chemoheterotrophic bacteria from the deep terrestrial subsurface

The metal resistance of 350 subsurface bacterial strains from two U.S. Department of Energy facilities, the Savannah River Site (SRS), South Carolina, and the Hanford site, Washington, was determined to assess the effect of metal toxicity on microorganisms in the deep terrestrial subsurface. Resistance was measured by growth inhibition around discs containing optimized amounts of Hg(II), Pb(II), and Cr(VI). A broad range of resistance levels was observed, with some strains of Arthrobacter spp. demonstrating exceptional tolerance. A higher level of resistance to Hg(II) and Pb(II) (P < 0.05) and a higher occurrence of multiple resistances suggested that metals more effectively influenced microbial evolution in subsurface sediments of the SRS than in those of the Hanford site. Common resistance to heavy metals suggests that toxic metals are unlikely to inhibit bioremediation in deep subsurface environments that are contaminated with mixed wastes.     

Reduction of cabbage root fly (Delia radicum) damage by protective discs

Foam-rubber, carpet-underlay discs placed in position around the base of the stem directly after transplanting were as effective as certain recommended insecticides in protecting brassica plants from damage by cabbage root fly (Delia radicum). The three factors that contributed to the overall effectiveness of the discs were that only half as many eggs were laid around plants protected with discs as around unprotected plants, that predatory ground beetles preferred to aggregate in the humid microhabitat beneath the discs and consequently ate proportionally more fly eggs/larvae and that the discs acted as a mulch, conserving water around the roots of the plants, and thereby permitting them to tolerate greater amounts of damage. Application of the repellent benzyl benzoate to the discs improved effectiveness but 12 other insect repellents and two sticky compounds gave no improvement and most were phytotoxic. Root drenches of the insecticide chlorfenvinphos were repellent, reducing oviposition by approximately 25%. The costs of protecting brassica transplants with either discs or insecticide were similar and discs thus appear to be a reasonable method of cabbage root fly control for small-scale use.     

Comparison of Different Agar Diffusion Methods for the Detection of Antimicrobial Residues in Slaughter Animals

The different agar diffusion methods were compared using antibiotic and sulphonamide-impregnated filter-paper discs and the kidneys of healthy and emergency-slaughtered pigs and cows after slaughter. No method used seemed to be sensitive to all antimicrobial drugs preimpregnated onto discs. Tetracycline yielded a greater zone of inhibition at pH 6 than at pH 8 and aminoglycosides, erythromycin, polymyxin B and lin cornycin at pH 8 than at pH 6. It therefore seems necessary to use different pHs (6 and 8). The addition of trimethoprim to the medium is necessary for the detection of sulphonamides. Bacillus subtilis BGA used as the test organism was more sensitive to sulphonamides on the “Test agar for the inhibitor test” containing trimethoprim than on the “Iso-Sensitest agar” also containing trimethoprim. The addition of trimethoprim to “Test agar for the inhibitor test” is recommended at pH 8 but not at pH 6 because false-positive cases (with inhibition zones > 2 mm) were observed at pH 6 with trimethoprim on the kidneys of healthy pigs.     

Inhibition of phytoalexin synthesis in arachidonic Acid-stressed potato tissue by inhibitors of lipoxygenase and cyanide-resistant respiration

Arachidonic acid-stressed potato tuber discs synthesized the phytoalexin rishitin. This synthesis was inhibited by salicylhydroxamic acid (SHAM), and to a lesser extent by tetraethylthiuram disulfide (disulfiram). Disulfiram was less effective apparently because it was inactivated in the tuber discs. Disulfiram and SHAM both inhibited cyanide-resistant respiration of whole potato discs and lipoxygenase extracted from these discs. When low disulfiram concentrations were used, the lipoxygenase inhibition was quickly overcome, again because the disulfiram apparently was inactivated by oxidation.     

Increased sensitivity of photosynthesis to antimycin A induced by inactivation of the chloroplast ndhB gene. Evidence for a participation of the NADH-dehydrogenase complex to cyclic electron flow around photosystem I

Tobacco (Nicotiana tabacum var Petit Havana) ndhB-inactivated mutants (ndhB-) obtained by plastid transformation (E.M. Horvath, S.O. Peter, T. Jo?t, D. Rumeau, L. Cournac, G.V. Horvath, T.A. Kavanagh, C. Sch?fer, G. Peltier, P. MedgyesyHorvath [2000] Plant Physiol 123: 1337-1350) were used to study the role of the NADH-dehydrogenase complex (NDH) during photosynthesis and particularly the involvement of this complex in cyclic electron flow around photosystem I (PSI). Photosynthetic activity was determined on leaf discs by measuring CO2 exchange and chlorophyll fluorescence quenchings during a dark-to-light transition. In the absence of treatment, both non-photochemical and photochemical fluorescence quenchings were similar in ndhB- and wild type (WT). When leaf discs were treated with 5 microM antimycin A, an inhibitor of cyclic electron flow around PSI, both quenchings were strongly affected. At steady state, maximum photosynthetic electron transport activity was inhibited by 20% in WT and by 50% in ndhB-. Under non-photorespiratory conditions (2% O2, 2,500 microL x L(-1) CO2), antimycin A had no effect on photosynthetic activity of WT, whereas a 30% inhibition was observed both on quantum yield of photosynthesis assayed by chlorophyll fluorescence and on CO2 assimilation in ndhB-. The effect of antimycin A on ndhB- could not be mimicked by myxothiazol, an inhibitor of the mitochondrial cytochrome bc1 complex, therefore showing that it is not related to an inhibition of the mitochondrial electron transport chain but rather to an inhibition of cyclic electron flow around PSI. We conclude to the existence of two different pathways of cyclic electron flow operating around PSI in higher plant chloroplasts. One of these pathways, sensitive to antimycin A, probably involves ferredoxin plastoquinone reductase, whereas the other involves the NDH complex. The absence of visible phenotype in ndhB- plants under normal conditions is explained by the complement of these two pathways in the supply of extra-ATP for photosynthesis.     

Ultraviolet-B Radiation-induced Inhibition of Leaf Expansion and Promotion of Anthocyanin Production: Lack of Involvement of the Low Irradiance Phytochrome System

Leaf discs from expanding leaves of Rumex patientia L. were exposed to 7 hours of visible plus different levels of ultraviolet radiation in the 290 to 315 nm waveband (UV-B) and then placed in darkness. Leaf disc expansion was reduced and anthocyanin production was increased in discs exposed to moderate or high levels of UV-B radiation when compared to control discs. The possibility that the inhibition of leaf expansion by UV-B radiation might be at least partially phytochrome-mediated was examined by giving discs brief red or far red irradiation following exposure to UV-B radiation. Brief red radiation (R) following treatment with moderate or high UV-B radiation did not alter the pattern of growth or anthocyanin production compared to discs placed in darkness following UV-B treatment. However, a posttreatment with far red radiation (FR) reduced the growth of discs subjected previously to either moderate UV-B or no UV-B irradiation to the level of growth of discs given high UV-B. FR posttreatment also decreased anthocyanin production in discs in moderate and high UV-B treatments. Effects of FR and UV-B radiation apparently do not involve the same mechanism. This was demonstrated by experiments in which FR following the UV-B treatments was in turn followed by R, which reversed the effects of the FR but did not alter the growth inhibition or increased anthocyanin production induced by moderate or high levels of UV-B radiation.     

Autoinhibition of Ethylene Production in Citrus Peel Discs : SUPPRESSION OF 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID SYNTHESIS

Wound ethylene formation induced in flavede tissue of citrus fruit (Citrus paradisi MacFad. cv. Ruby Red) by slicing was almost completely inhibited by exogenous ethylene. The inhibition lasted for at least 6 hours after removal of exogenous ethylene and was then gradually relieved. The extent of inhibition was dependent upon the concentration of ethylene (1 to 10 microliters/liter) and the duration of treatment. The increase in wound ethylene production in control discs was paralleled by an increase in 1-aminocyclopropane-1-carboxylic acid (AAC) content, whereas in ethylene-treated discs there was little increase in ACC content. Application of ACC completely restored ethylene production in ethylene-pretreated discs, indicating that the conversion of ACC to ethylene is not impaired by the presence of ethylene. Thus, autoinhibition of ethylene synthesis was exerted by reducing the availability of ACC. Ethylene treatment resulted in a decrease in extractable ACC synthase activity, but this decrease was too small to account for the marked inhibition of ACC formation. The data indicate that autoinhibition of ethylene production in citrus flavede discs results from suppression of ACC formation through repression of the synthesis of ACC synthase and inhibition of its activity.     

Mathematical model for the control and standardization of discs]

Analysis of tetracycline and erythromycin diagnostic discs manufactured in the USSR showed their conformity with the requirements of the USA Federal Register. Comparison of the antibiotic amounts extracting and diffusing from the discs showed that as an average 92 and 90 per cent of the extracted amounts of erythromycin and tetracycline respectively diffused into the agar. Subsequently, it is desirable that on control testing of the disc quality both the similarity of the antibiotic content in the discs and the real amount of the antibiotic diffusing into the agar should be considered. A statistically reliable correlation between the values of the growth inhibition zones around the discs with definite and constant amounts of erythromycin, tetracycline or oxacillin and different resistance levels for every staphylococcal strain was found. On the basis of such a control system it is possible to divide the staphyloccal strains into the groups with high, low and intermediate resistance levels to the above antibiotics. However, it is not possible to use such data for accurate calculation of the value of the minimum inhibitory concentration of unknown strains because of a high value of the main error.     

Susceptibility of Pseudomonas aeruginosa to Gentamicin Sulfate In Vitro: Lack of Correlation Between Disc Diffusion and Broth Dilution Sensitivity Data

Seventy-eight of 420 clinical isolates of Pseudomonas aeruginosa yielded zones of inhibition of less than 12 mm in diameter around 10-mug discs of gentamicin sulfate when tested by the standardized Bauer-Kirby disc diffusion method. Of 153 strains chosen from these isolates, one strain (0.65%) required 25 mug of gentamicin per ml for inhibition; the remainder (99.35%) were inhibited by 6 mug/ml or less of the antibiotic. It is recommended that those isolates of P. aeruginosa that yield zones of inhibition less than 12 mm in diameter be disc susceptibility-tested once more; those isolates that give zones of inhibition of less than 12 mm upon repeated examination should then be subjected to the broth dilution test before they are designated as sensitive or resistant to gentamicin.     

Chromium-induced inhibition of ethylene evolution in bean (Phaseolus vulgaris) leaves

The influence of chromium concentration on ethylene production in bean plants ( Phaseolus vulgaris L. cv. Contender) was investigated. A Cr ion-induced inhibition of ethylene synthesis from endogenous 1-aminocyclopropane-1-carboxylic acid (ACC) was observed within both leaf discs floated on 2 m M CrO²⁻₄ or Cr³⁺ and leaf discs from plants cultured in nutrient solutions containing 10, 20 or 40 μ M CrO²⁻₄. However, Cr ions supplied either to plants with the nutrient solution or to discs with the incubation medium rather increased the conversion of exogenous ACC to ethylene. Primary leaves of plants exposed to CrO²⁻₄-containing nutrient solutions showed a statistically insignificant decrease of ACC-synthase activity. In the trifoliolate leaves of plants exposed to 10 μ M CrO²⁻₄, in which a significant decrease of ethylene production from endogenous ACC was observed, a substantial increase of ACC synthase was found. These results indicate that Cr ion-induced inhibition of ethylene production is not due to a breakdown of membrane integrity, which is necessary for ethylene forming enzyme activity, but caused by metabolic alterations leading to decreased ACC availability. Chromium ions may act by inhibiting ACC synthase activity or by diverting a metabolic step prior to the ACC synthase catalyzed reaction.     

Effect of cytochalasin B on the evagination in vitro of leg imaginal discs

Cytochalasin B (1 μg/ml) completely inhibited the evagination of isolated leg imaginal discs cultured in vitro in a synthetic medium (ME) containing α-ecdysone (3 μg/ml). In discs precultured for 6 hr in medium ME without the drug, then transferred to cytochalasin B-containing medium, continuation of evagination was stopped immediately. The inhibition of evagination was completely reversible, provided pretreatment with cytochalasin B did not exceed 8 hr. Results are discussed in view of what is known on the effect of cytochalasin B on other developmental systems. Findings are compatible with the primary action of the drug being an alteration of cell surface properties, thus bringing to light the importance of these properties in the course of normal imaginal disc evagination.     

Soil fungistasis: elevation of the exogenous carbon and nitrogen requirements for spore germination by fungistatic volatiles in soils.

Axenic, washed conidia of Fusarium solani f. sp. phaseoli, Aspergillus flavus, and Verticillium albo-atrum were placed on washed Difco purified agar discs along with an inorganic salt solution containing various levels of carbon and nitrogen substrates. These discs were exposed to volatiles from six soils (pH 5.1-8.6). Fusarium solani macroconidial germination was inhibited mostly by volatiles from soils of pH 5.1, 6.1, 7.0, and 7.5, but high levels of glucose and NH4Cl reversed this inhibition, raising germination to that of no-soil, no-carbon or nitrogen controls. Conidial germination of A. flavus was inhibited mainly by volatiles from high pH (7.0, 7.8, and 8.6) soils, and increased levels of glucose plus an amino acid mixture nullified this inhibition. Volatiles from soils of pH 5.1, 6.1, and 7.5 stimulated A. flavus conidial germination. Assays after the removal of CO2 from the air above soil of pH 5.1 demonstrated that volatiles inhibitory to A. flavus were produced by this soil. Assays indicated that a KOH-soluble compound was a fungistatic soil volatile to F. solani macroconidial germination. The nullification by carbon and nitrogen substrates of F. solani and A. flavus inhibition caused by soil volatiles parallels that for soil fungistasis. Conidial germination of V. albo-atrum was markedly stimulated by volatiles in all soils tested, and was not affected by removal of CO2. Inhibitory soil volatiles may increase the nutritional requirements for spore germination of certain fungi.